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Compatibility Assessment of a Model Monoclonal Antibody Formulation in Glass and in Blow-Fill-Seal (BFS) Plastic Vial Delivery Formats

Dipesh Shah, Ph.D. Senior Development Engineer Advanced Delivery Technologies, Woodstock, Illinois

©2014 Catalent Pharma Solutions. All rights reserved

Presentation Outline

1. Advanced Aseptic Delivery Technology: Blow-Fill-Seal

2. General Requirements for Container Qualification and examples of Protein-Container Interactions

3. Case – Study: Compatibility assessment of a model monoclonal antibody formulation with AdvaseptTM and Glass vial

1: Advanced Aseptic Processing: Blow Fill Seal

Quality/Sterility Assurance

Minimizing Particulates

Stability/ Compatibility

Reduce risk factors for sterility challenges through automation

Elimination of glass particles and delamination with significant reduction in foreign particulates

Medical grade polypropylene resin for excellent chemical and physical properties

Advanced Aseptic Filling Technology

Why Glass Free?

Safety/Reduced Product Loss

Plastic construction reduces risk of breakage and simplifies opening

Advanced Delivery Technologies for Parenterals to Mitigate Risk Associated with Aseptic Processing

Minimize Risk: Simplify the Process, Reduce Variables

In just 15 seconds, the container is formed, filled and

sealed in ISO 5 aseptic conditions

Minimizing Variables and the Footprint in Aseptic Manufacturing

Traditional Glass Vial Filling

Vials Stoppers

Blow Fill Seal

Sterilized Stopper

insertion for vials

Form

Fill

Insertion

Seal

Prequalified Resin

Downstream equipment

Utilizing the automated aseptic design of BFS, the technology eliminates traditional manufacturing steps, reduces the required controlled space and

decreases the risk of contamination associated with traditional aseptic practices

Glass Vial Filling

Large controlled space

Glass management and process control

Blow Fill Seal

Class A space 10 sq. ft

Sterile container is formed at the time of fill

Leveraging Advanced Aseptic Process Blow Fill Seal Technology

Reference: Verjans, B. Reed, C. (2012). “Assessing Filling Technologies for Contamination Risk.” Biopharm International. 25(3), pp. 46-58.

The BFS process minimizes the risk of contamination by reducing particles, process steps & human interaction

Potential risk of contamination by filling technology based on air quality and exposure time

2: General Requirements for Container Qualification and Examples of Protein-Container Interactions

General Requirements for Container Qualification

Safety –

• Are we maintaining sterility?

• What are we leaching?

Efficacy –

• Are we adequately protecting the dose?

• Is there binding -- adsorption/absorption?

• Are we delivering the dose?

Container Interactions with Protein Therapeutics

Discussion Topics

1.What are the product development concerns for container interactions with protein products?

2.What testing is needed to understand container interactions with protein therapeutics?

Liquid Formulation Containers (Injectables)– Potential Materials

Enhanced System Container Materials Closure Materials

Vials or Ampoules

(SVP)

Glass/AdvaseptTM Vials with Butyl

rubber/Teflon coated butyl

rubber

Prefilled syringes

with plungers

Glass or Plastic

Staked needle or luer lock

(plastic or rubber)

Butyl rubber

Teflon coated butyl rubber

IV Infusion (LVP) Glass/Flexible Bags with

spike ports/AdvaseptTM

Polyethylene

General Concerns for Protein-Container Interactions

Container Concerns Product concerns

• Protein adsorption • Protein loss

• Headspace • Agitation causes protein aggregation

(AdvaseptTM Advantage)

• Organic leachables (eg.

Silicone, DEHP)

• Toxicity of leachable

• Protein aggregation/Immunogenicity

• *Trace metals (eg. Iron,

Tungsten, Aluminum)

• Eg. Iron – Iron Leachable catalyzed protein

degradation reactions

• Eg. Tungsten – Protein Oxidation

(AdvaseptTM Advantage)

• Eg. Aluminum – Aluminum Phosphate

resulted in visible particles (AdvaseptTM

Advantage)

*www.pqri.org/workshops/podp11/pdfs/markovic.pdf , “Regulatory perspective safety qualification of extractable and leachable,”

Feb. 22-23, 2011, Ingrid Markovic, Ph.D.

Challenges to fill Biologics with Blow-Fill- Seal (BFS)

Challenges:

1. Impact of elevated temperature of the molded plastic during filling on the stability of the biologic formulation

2. Impact of gas permeation from semi-permeable BFS plastic container on degradation of biologics (for eg. Oxidation)

3. Impact of potential leachables from BFS processed plastic container system on biologic stability and safety of the formulation.

Temperature Profile of Resin and Container

Temperature Profile of solution

Conclusion: The molded plastic process was optimized by Catalent to reduce the temperature of the solution (0.5 mL fill) close to 40°C at time of fill and the assumption is that the temperature of the solution would fall steadily after the units come out of the fill suite and packaged under ambient conditions.

Oxygen and Carbon Dioxide Gas Permeation Data

Conclusion: Carbon Dioxide has higher permeation rate relative to Oxygen because Carbon Dioxide partitions into the container to a greater extent relative to Oxygen.

0.0E+00

2.5E-02

5.0E-02

7.5E-02

1.0E-01

0 10 20 30 40

cc*

mm

/(cm

2*

day)

Temperature (C)

Gas Permeation Rate vs Temperature (Normalized)

Oxygen

Carbon

Dioxide

Water- Vapor Transmission Rates

0.0E+00

1.0E-05

2.0E-05

3.0E-05

4.0E-05

5.0E-05

5 10 15 20 25 30 35 40

g*m

m/(

mm

2*day)

Temperature (C)

Water Vapor rate (normalized) vs Temperature (C)

Conclusion: Higher the Temperature, greater the water vapor permeation rates

Case – Study: Compatibility assessment of a model monoclonal antibody formulation with AdvaseptTM and Glass container system

Monoclonal Antibody Formulation Recipe + Preparation

.

Table 1: Model mAb Formulation

Components Amount

(mg/mL)

Model mAb ( Mol.

Wt. 144 kDa)

10

Polysorbate 80 0.7

Sodium Citrate 6.5

Sodium Chloride 9.0

pH 6.5

The monoclonal antibody formulation was filtered with a 0.2 micron Nalgene filter unit

and filled in glass vials into which uncoated stoppers were placed. The mAb formulation

was filtered into a sterile bag and shipped to Blow-Fill-Seal (BFS) manufacturing facility.

The formulation was aseptically transferred in BFS AdvaseptTM

stoppered vials.

mAb Compatibility Assessments (Glass/Advasept Vial formats)

Parameter Method Target Range pH USP 791 6.5 + 0.3

Appearance Visual Inspection Report Results

Potency

UV (280 nm) T=0 + 10%

Activity Report EC50

Stability

Size-Exclusion Chromatography (SEC)

Report % Monomer and % High and low molecular weights

Nanoparticle Tracking Analysis (NTA)5

Report sub-micron particle size analysis

Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis

(SDS-PAGE)

Report % Area (HC +LC)

Capillary Isoelectric focusing (cIEF) pI (% of each peak)

Peptide Mapping2 % Chemical Modification

Leachables3

Polar Leachables (HPLC-UV) Report Irganox 1010 levels

Semi-Volatile Leachables (GC-MS) Report Aromatic Hydrocarbon levels

Volatile Leachables GC-FID Report Volatile Leachable levels4

Metals (ICP-MS) Report all metals except Na, I

Bacterial Endotoxin1 USP 85 Report Results

Bioburden1 USP 61 <10 CFU/mL 1 only performed at 0 time-interval

2 Only performed at 4 month time-interval 3 Only performed at 9M time-interval

4 Isopropyl alcohol, Methyl Ethyl Ketone, Trichloroethylene, Hexane, 2-methyl pentane, 3-methyl pentane, Methylcyclopentane, Cyclohexane and Heptane

5 Performed on 12 M samples

Results 1 -Potency

Potency:

a) UV: The UV data indicates no apparent change in potency for both vial formats

upon pre and post-fill and upon storage to 9 months at 5°C were within target range.

b) Complement Dependent Cytotoxic Assay: The potency was determined using responsive cell line in a complement dependent cytotoxic assay using a fluorescence read out. Comparison of a dilution series with standard, formulation in glass and the AdvaseptTM vial were generated (Figure 1).

Figure 1: Dose Response Curve for model mAb formulation

x axis

0.001 0.01 0.1 1 10

0

1000

2000

3000

Graph#1

y = ( (A - D)/(1 + (x/C)^B ) ) + D: A B C D R^2

Advasept 2 (Advasept 2: Conc vs MeanValue) 3574.447 1.618 0.373 247.764 0.996

Glass vial 2 (Glass Vial 2: Conc vs MeanValue) 3511.099 1.729 0.482 325.305 0.998

Advasept 1 (Advasept 1: Conc vs MeanValue) 3534.978 1.711 0.391 320.817 0.998

Glass vial 1 (Glass vial 1: Conc vs MeanValue) 3568.287 1.728 0.46 338.779 0.999

Conclusions: The activity data shows comparable potency values between the Glass and AdvaseptTM

vials upon storage to 24M/5°C (All data not shown).

Result 3 -Nanoparticle Tracking Analysis (NTA)

Particles AdvaseptTM

Glass Vial

D50 126 + 17.5 94 + 10.1 nm

D90 205 + 35 nm 144 + 13.4 nm

Total

Concentration

(particles/frame)

13.95 + 3.2 13.47 + 3.0

Total

Concentration

(particles/mL)

2.83e8 + 6.5e

7 2.74e

8 + 6.1e

7

Mean sub-micron particle count data

(Nanoparticle Tracking Analysis)

for Monoclonal antibody formulations stored in

Glass and AdvaseptTM

vials for 12M/5°C.

Conclusions: Slightly higher size mode of aggregates were observed in AdvaseptTM than in the glass vials, however, the total number of particles are statistically comparable between the two container closure systems.

Result 2 SEC-UV Aggregation Data

Conclusion: Stability data indicates higher levels of high

molecular weight species in Glass relative to AdvaseptTM vial.

0

1

2

3

4

5

6

0 6 12 18 24

% H

igh

Mole

cu

lar

Weig

ht

Sp

ecie

s

Months

SEC-UV - Aggregation data (5C)

Results 4-Peptide Mapping Results

Type Sample % Modification

Oxidation Glass Vial 10.8

AdvaseptTM Vial 4.5

Deamidation Glass Vial 7.2

AdvaseptTM Vial 7.6

Conclusions: The oxidation levels were higher in Glass relative

to AdvaseptTM vials and could be attributed to mAb’s surface

interaction with glass surface causing more oxidation than in

plastic AdvaseptTM vials.

Procedure: Aged Glass and AdvaseptTM

vials (4M/5°C) were subjected to peptide

mapping by initially denaturing and reduction of the mAb with DTT, alkylation with

Iodoacetamide, followed by clean-up on a column, and digestion with Trypsin and

ASP-N. The peptides were separated on a UPLC column and UV and MS detectors

were employed. (Chromatograms provided in the appendix section).

Results 5 (pH, Appearance, cIEF and SDS-PAGE)

All other stability test results for parameters listed below were comparable between Glass and AdvaseptTM vial formats.

1. pH

2. Appearance

3. cIEF

4. SDS-PAGE

Bioburden results performed at t=0 for both vial configurations

were below <10 CFU/mL.

Results 6 (Leachable Data Analysis)

The monoclonal antibody formulation samples stored for 9M at 5°C were analyzed for various leachables listed below:

1.Volatile: GC-FID

2.Semi-Volatile: GC-MS

3.Polar: HPLC-UV

4.Metal leachables: ICP-MS

Results: The leachable data indicates comparable leachable profile for monoclonal antibody formulations stored in Glass and AdvaseptTM vials except for Isopropyl alcohol. The Isopropyl alcohol content was determined to be higher in Glass vial (14 mg/mL) relative to AdvaseptTM vial (1.2 mg/mL) and was attributed to cleaning procedure used for glass vials prior filling of the monoclonal antibody formulation.

Conclusions

1. This study confirms compatibility of monoclonal antibody formulation in glass and AdvaseptTM vial demonstrating plastic BFS vial suitability for protein therapeutics.

2. Stability data indicates higher levels of high molecular weight species in Glass relative to AdvaseptTM vial.

3. Affinity data indicates comparable potency levels in Glass and AdvaseptTM vial.

4. Leachable data indicates comparable leachable profiles in Glass and AdvaseptTM vial.

5. No significant differences were identified between Glass and AdvaseptTM vials over the 9 months analyzed to date.

ACKNOWLEDGEMENTS

Catalent- Woodstock Catalent – Kansas City

1. Waiken Wong, Ph.D. 1. Vincy Abraham, Ph.D.

2. Bill Hartzel

3. Natasha Hults Catalent - RTP

4. Kay Schmidt 1. Thomas Luntz

2. Courtney Jones

Catalent – Madison 3. James Mclean

1. Gregory Bleck, Ph.D. 4. Vicki Wards

2. Ian J. Collins, Ph.D.

3. Process Development Team

QUESTIONS ???

APPENDIX

cIEF Analysis

cIEF Chromatogram Time 0

a) Glass Vial b) AdvaseptTM

vial

Magnified Chromatograms Time 0

a) Glass Vial b) AdvaseptTM

vial

The chromatograms showed no comparable charge distribution between the Glass and AdvaseptTM

vial.

Impact of Elevated Temperature of the Molded Plastic during filling on the stability of biologic formulation

T=0 data

Parameters Bulk Solution Glass Vial AdvaseptTM Vial

pH 6.4 6.6 6.6

Potency (UV) (mg/mL)

10.8 10.8 10.9

SEC 2.2% High Mol. Wt. 97.8% Monomer

2.1% High Mol. Wt 97.9% Monomer

2.1% High Mol. Wt. 97.9% Monomer

SDS Similar Profile (Reduced/Non Reduced) Figures – Appendix section

cIEF Major PIs (%)* pI Range 9.38-9.65 (7

peaks)

Major PIs (%) pI Range 9.42-9.74 (8

peaks)

Major PIs (%) pI Range 9.42-9.74 (8

peaks)

Conclusion: Data from stability indicating parameters between the bulk mAb, glass and AdvseptTM vial

drug product indicates that the BFS process is amenable to Biologics.

Peak pI % Area

1 9.38 11.0

2 9.40 14.0

3 9.45 11.9

4 9.50 34.0

5 9.56 3.49

6 9.59 20.7

7 9.65 5.0

Peak pI % Area

1 9.42 15.9

2 9.44 3.2

3 9.49 18.8

4 9.54 31.8

5 9.58 6.2

6 9.62 18.9

7 9.67 5.1

8 9.74 0.1

Peak pI % Area

1 9.42 15.8

2 9.44 3.1

3 9.49 18.6

4 9.54 31.9

5 9.58 6.2

6 9.62 19.1

7 9.67 5.3

8 9.74 0.2

*analysis occurred at separate time using different cIEF cartridge which may explain slight differences and possible poor resolution of peak 8 which is only 0.1% of the total area. Peak profiles look similar.

cIEF (peaks)

SDS – PAGE Analysis

SDS-PAGE (Non-Reduced)

Lane 5 – Glass Vial

Lane 6 – AdvaseptTM

Vial

SDS-PAGE (Reduced)

Lane 5 – Glass Vial

Lane 6 – AdvaseptTM

Vial

Magnified UPLC/UV Chromatograms of Trypsin Digested Monoclonal Antibody

AU

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Magnified UPLC/UV Chromatograms of ASP-N Digested Monoclonal Antibody

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Let us meet again..

We welcome you all to our future conferences of OMICS International

2nd International Conference and Expo on

Parenterals and Injectables

On

October 24-26, 2016 at Istanbul, Turkey http://parenterals-

injectables.pharmaceuticalconferences.com/